Estrogen hormone action within target tissues involves the interaction of the hormonal substance with two different receptor proteins termed ER-alpha and ER-beta. The specificity and responsiveness of tissues to hormonal stimulation are governed in most part by the presence and biochemical action of these receptor proteins. Mouse lines recessive for functional ER-alpha (ERKO) and ER-beta (BERKO) receptor signaling systems anbd the recent generation of double ERKO lines (DERKO) having neitehr functional ER-alpha or ER-beta, have been produced and characterization of phenotypes in the reproductive, cardiovascular, immune, skeletal and neuroendocrine systems are currently underway. Initial characterization shows different phenotypes in the BERKO than the ERKO; with the major effect in the ovary resulting in subfertility and faulty folliculogenesis. Because of this phenotype we are evaluating whether mutations or polymorphism in the ER-beta gene may correlate with clincal cases of female infertility. Double ERKO females have a unique ovarian phenotype resulting in infertility, no folliculogensis but laso trandifferentiation of granulosa cells to sertoli cells. Such results indicate that both ER-alpha and ER-beta are needed to maintain proper granulosa cell differentiation. BERKO male mice are fertile in contrast to ERKO males which are infertile. Nuclear localization of ER-alpha and its activity was increased by exposure to growth factors, suggesting a coupling of these two signaling pathways. Use of the alpha-ERKO mice has supported the coupling as an in vivo mechanism, since EGF and IGF-1 treatments did not stimulate uterine DNA synthesis in the absence of a functional estrogen receptor. Characterization of the EGF and IGF-1 signaling pathway in the mice indicates EGF and IGF-1 receptor levels and functionally are not altered compared to controls including the phosphorylation of IRS-1 a pivitol component in IGF signaling. ERKO were used as an experimental model system for evaluating the role of ER-alpha in normal physiological endocrine function and hormonal carcinogenesis. Since ERKO mice have no mammary gland development, a test for the role of estrogen receptor in development of mammary tumors was developed. A cross of ERKO with a mammary sensitive mouse line (WNT-1) expressing the WNT-1 oncogene was produced. These data indicate that the WNT-1 oncogene does not require functional estrogen receptor to produce an epithelial cell specific mitogenic response. Additionally, these animals do develop tumors at approximately half the level as WT. These findings would oppose one earlier hypothesis that mammary tumors develop from hormone responsive tissues. ERKO male infertility was shown to be dependent on a progressive lesion of testicular semiferous tubule dysmorphogenesis resulting in a loss of germ cells and decreased sperm counts. ERKO male infertility was rescued using sperm transplantation techniques indicating the need for ER-alpha was not in the germ cell but in the somatic cells of the male reproductive tract. The findings also clearly demonstrate a requirement for estrogen receptor action in normal male reproductive physiology. As a means of extending the ERKO model evaluating the mechanism of environmental estrogen action ERKO mice were treated with genistein which also acts as a tyrosine kinase inhibitor and shows that uterine growth was not stimulated indicating genistein acts through the estrogen receptor for this response. However, suppression of lutenizing hormone (LH) was shown to be as effective in ERKO as WT mice indicating this estrogen response by genistein does not require the receptor as previously thought. Differences in gene responses to hormonal xenobiotics were also seen in some in vitro gene regulation studies. Using transactivation assay system trichlorobiphenol was shown to be more active than estradiol on the lactoferrin gene. Use of this estrogen responsive yeast system also tested the xenobiotic estrogen synergy concept and showed that using a variety of genes that synergy was not apparent and had no universal application to hormone responsive systems.